The growth hormone from man and from the common domestic animals are proteins of approximately 191 amino acids, synthesized and secreted from the anterior lope of the pituitary gland. Human growth hormone consists of 191 amino acids with the sequence FPTIPLSRLFDNAMLRAHRLHQLAFDTYQEFEEAYIPKEQKYSFLQNPQTSLCFSESIPTPSN REETQQKSNLELLRISLLLIQSWLEPVQFLRSVFANSLVYGASDSNVYDLLKDLEEGIQTLMG RLEDGSPRTGQIFKQTYSKFDTNSHNDDALLKNYGLLYCFRKDMDKVETFLRIVQCRSVEG SCGF, SEQ ID No. 1.
Growth hormone is a key hormone involved in the regulation of not only somatic growth, but also in the regulation of metabolism of proteins, carbohydrates and lipids. The major effect of growth hormone is to promote growth. The organ systems affected by growth hormone include the skeleton, connective tissue, muscles, and viscera such as liver, intestine, and kidneys.
The metabolic effect of growth hormones is mediated by Insulin-like Growth Factor-I (IGF-I). IGF-I is primarily produced in the liver and it circulates in the plasma bound to specific binding proteins, the most important of which is Insulin-like Protein Binding Protein-3 (IGFBP-3). Growth hormone appears to exert its activity through two distinct mechanisms. In the post-prandial period, the effect is mediated through IGF-1 and result is glucose uptake, lipogenesis and proteinogenesis. When fasting, i.e. when the blood glucose is low, the growth hormone secretion is boosted, where it increases lipoxidation. It is thus noteworthy that growth hormone both in the post-prandial period and in fasting periods acts to preserve and build up proteins.
Growth hormone is used to treat growth hormone deficiency, e.g. various forms of short stature. Unfortunately, growth hormone has a relative short half life which means that patients receiving growth hormone treatment typically need daily growth hormone administration. Growth hormone being a protein, the administration form is injections which represents a daily inconvenience to the patients.
In an attempt to lower the administration frequency, growth hormone in a sustained release formulation has been marketed under the trade name Nutropin Depot®. Another approach to lower the administration frequency is to manipulate the growth hormone molecule to increase the half life. Well-known ways to increase half life of proteins include PEGylation (U.S. Pat. No. 4,179,337 and J. Biol. Chem. 271, 21969-21977 (1996)), which probably works by increasing the molecular size of the molecule to decrease renal clearance and by making the molecule less susceptible to protease break down.
The PEG moiety may be attached to PEG via different forms of bonds or linkages including amide bonds obtained by acylation of free amino groups. Alternatively, a functional group may be introduced into growth hormone, which group is reacted with a functionalised PEG moiety in a subsequent step. This approach often has the advantage of improved selectivity as the introduced functional group may be selected so that it is unique to growth hormone. U.S. Pat. No. 6,673,347 discloses that polymeric groups may be attached to proteins by means of an oxime bond. An oxime bond may be formed in a reaction between a carbonyl functionalised, such as an aldehyde functionalised growth hormone and an amineoxy functionalised PEG, or vice verse in a reaction between an amineoxy functionalised growth hormone and a carbonyl, such as an aldehyde functionalised PEG.
WO 97/39768 discloses that human growth hormone may be formulated at pH between 6.0 and 8.8.
J. Am. Chem. Soc. 117, 3893-3899 (1995) discloses that the rate of oxime bond formation has its maximum at pH fairly low pH, i.e. around or below 5.7.
The present invention aims at providing pharmaceutical compositions for pegylated growth hormone wherein the PEG is attached to the growth hormone via an oxime bond. Such formulations have improved or alternative properties compared to known formulations.